Confocal and multiphoton imaging of intracellular Ca²⁺

This chapter compares the imaging capabilities of a range of systems including multiphoton microscopy in regard to measurements of intracellular Ca<sup>2+</sup> within living cells. In particular, the excitation spectra of popular fluorescent Ca<sup>2+</sup> indicators are shown during 1P and 2P excitation. The strengths and limitations of the current indicators are discussed along with error analysis which highlights the value of matching the Ca<sup>2+</sup> affinity of the dye to a particular aspect of Ca<sup>2+</sup> signaling. Finally, the combined emission spectra of Ca<sup>2+</sup> and voltage sensitive dyes are compared to allow the choice of the optimum combination to allow simultaneous intracellular Ca<sup>2+</sup> and membrane voltage measurement

Tunable Pinning of Burst-Waves in Extended Systems with Discrete Sources

We study the dynamics of waves in a system of diffusively coupled discrete nonlinear sources. We show that the system exhibits burst waves which are periodic in a traveling-wave reference frame. We demonstrate that the burst waves are pinned if the diffusive coupling is below a critical value. When the coupling crosses the critical value the system undergoes a depinning instability via a saddle-node bifurcation, and the wave begins to move. We obtain the universal scaling for the mean wave velocity just above threshold.Comment: 4 pages, 5 figures, revte

Endoplasmic Reticulum Remodeling Tunes IP3-Dependent Ca2+ Release Sensitivity

The activation of vertebrate development at fertilization relies on IP3-dependent Ca2+ release, a pathway that is sensitized during oocyte maturation. This sensitization has been shown to correlate with the remodeling of the endoplasmic reticulum into large ER patches, however the mechanisms involved are not clear. Here we show that IP3 receptors within ER patches have a higher sensitivity to IP3 than those in the neighboring reticular ER. The lateral diffusion rate of IP3 receptors in both ER domains is similar, and ER patches dynamically fuse with reticular ER, arguing that IP3 receptors exchange freely between the two ER compartments. These results suggest that increasing the density of IP3 receptors through ER remodeling is sufficient to sensitize IP3-dependent Ca2+ release. Mathematical modeling supports this concept of ‘geometric sensitization’ of IP3 receptors as a population, and argues that it depends on enhanced Ca2+-dependent cooperativity at sub-threshold IP3 concentrations. This represents a novel mechanism of tuning the sensitivity of IP3 receptors through ER remodeling during meiosis

Data-Driven Modelling of the Inositol Trisphosphate Receptor (IPR) and its Role in Calcium-Induced Calcium Release (CICR)

We review the current state of the art of data-driven modelling of the inositol trisphosphate receptor (IPR). After explaining that the IPR plays a crucial role as a central regulator in calcium dynamics, several sources of relevant experimental data are introduced. Single ion channels are best studied by recording single-channel currents under different ligand concentrations via the patch-clamp technique. The particular relevance of modal gating, the spontaneous switching between different levels of channel activity that occur even at constant ligand concentrations, is highlighted. In order to investigate the interactions of IPRs, calcium release from small clusters of channels, so-called calcium puffs, can be used. We then present the mathematical framework common to all models based on single-channel data, aggregated continuous-time Markov models, and give a short review of statistical approaches for parameterising these models with experimental data. The process of building a Markov model that integrates various sources of experimental data is illustrated using two recent examples, the model by Ullah et al. and the “Park–Drive” model by Siekmann et al. (Biophys. J. 2012), the only models that account for all sources of data currently available. Finally, it is demonstrated that the essential features of the Park–Drive model in different models of calcium dynamics are preserved after reducing it to a two-state model that only accounts for the switching between the inactive “park” and the active “drive” modes. This highlights the fact that modal gating is the most important mechanism of ligand regulation in the IPR. It also emphasises that data-driven models of ion channels do not necessarily have to lead to detailed models but can be constructed so that relevant data is selected to represent ion channels at the appropriate level of complexity for a given application